Phytophthora spp are pathogenic agents from the family of Pythiaceae known to be involved in various plant diseases. Sudden oak death, soybean root rot, apple crown and collar rot, root rot, affecting American chestnuts, rhododendron, African violet, or strawberries, are just examples of diseases caused by this group of pathogens. Typically, plant diseases caused by Phytophthora are considerably difficult to control and often lead to the death of the plant. This pathogen is a widespread and an economic problem to growers around the world. For example, Phytophthora infestans was the infective agent of the potato that caused the Great Irish Famine between 1845 and 1849. Presently, in North America growers are still facing the ravage of this pathogenic agent. Many conventional management techniques such as rootstock selection and site modification to control Phytophthora spp have been mostly unreliable.
Crown and collar rot, caused by at least four Phytophthora spp. (Jeffers et al., Phytopathology 2:533-538 (1982)) is a widespread and economically serious problem of apples throughout the Northeast U.S. In New York State, this disease appears to be the most common biological cause of premature tree decline and death, and, in Pennsylvania, many growers have abandoned the horticulturally-desirable MM 106 rootstock because of high crown rot incidence or its perceived threat. Stem and root rot of soybeans caused by Phytophthora sojae Kaufmann and Gerdemann (also denoted Phytophthora megasperma forma specialis glycinea) is also a widespread and serious problem. Because no single approach to the control of Phytophthora crown rot has proved reliable, growers have been advised to adopt an integrated or additive disease management strategy, utilizing a combination of site selection, site modification, rootstock selection, and chemical treatments where appropriate.
Pythium, like others in the family Pythiaceae, are usually characterized by their production of coenocytic hyphae, hyphae without septations. These are commonly called water molds. Pythium damping off is a very common problem in fields and greenhouses, where the organism kills newly emerged seedlings. This disease complex usually involves other pathogens such as Phytophthora and Rhizoctonia. Pythium wilt is caused by zoospore infection of older plants leading to biotrophic infections that become necrotrophic in response to colonization/reinfection pressures or environmental stress, leading to minor or severe wilting caused by impeded root functioning. See Jarvis, W. R., “Managing Diseases in Greenhouse Crops,” APS Press, St. Paul, Minn. (1992); Bagnall, R., “Control of Pythium Wilt and Root Rot of Hydroponically Grown Lettuce by Means of Chemical Treatment of the Nutrient Solution,” M. Sc Thesis, University of Pretoria, Pretoria, South Africa (2007); Plaats-Niterink A J van der, “Monograph of the Genus Pythium,” Studies in Mycology 21:1-242 (1981); Levesque et al., “Molecular Phylogeny and Taxonomy of the Genus Pythium,” Mycological Research 108:1363-1383 (2004); Jarvis, W. R., “Managing Diseases in Greenhouse Crops,” APS Press, St. Paul, Minn. (1992); Owen-Going, T. N., “Etiology and Epidemiology of Pythium Root Rot in Bell Pepper (Capsicum annuum L.) in Commercial-Scale and Small-Scale Hydroponic Systems,” M.Sc. thesis, University of Guelph, Guelph, Ontario (2002); Owen-Going et al., “Relationships of Pythium Isolates and Sweet Pepper Plants in Single-Plant Hydroponic Units,” Canadian Journal of Plant Pathology 25:155-167 (2003); Owen-Going, T. N., “Quantitative Investigations of Phenolic Compounds Associated With Root Rot of Hydroponic Pepper (Capsicum annuum L. Caused by Pythium aphanidermatum, (Edson) Fitzp. Ph.D. Thesis, University of Guelph, Guelph, Ontario (2005).
Many Pythium species, along with their close relatives, Phytophthora species are plant pathogens of economic importance in agriculture. Pythium spp. tend to be very generalistic and unspecific in their host range. They infect a large range of hosts, while Phytophthora spp. are generally more host-specific. For this reason, Pythium spp. are more devastating in the root rot they cause in crops, because crop rotation alone will often not eradicate the pathogen (nor will fallowing the field, as Pythium spp. are also good saprotrophs, and will survive for a long time on decaying plant matter).
Fusarium is a large genus of filamentous fungi widely distributed in soil and in association with plants. Most species are harmless saprophytes and are relatively abundant members of the soil microbial community. Some species produce mycotoxins in cereal crops that can affect human and animal health if they enter the food chain. The main toxins produced by these Fusarium species are fumonisins and trichothecenes. The genus includes a number of economically important plant pathogenic species. See Priest and Campbell, “Brewing Microbiology,” 3rd edition., ISBN 0-306-47288-0; Walsh et al., “Spectrum of Mycoses,” In: Baron's Medical Microbiology (Baron S et al, eds.), 4th ed., Univ of Texas Medical Branch. (via NCBI Bookshelf) ISBN 0-9631172-1-1 (1996); Howard, D H, “Pathogenic Fungi in Humans and Animals,” 2nd ed., Marcel Dekker. (via Google Books) ISBN 0-8247-0683-8 (2003); Van der Walta et al., “Fusarium Populations in the Household Food Gardens of a Peri-Urban Community,” South African Journal of Science 103 (2007); World Health Organization (1999-09-01), “Toxic Effects of Mycotoxins in Humans” (2007); Drug Policy Alliance, “Repeating Mistakes of the Past: Another Mycoherbicide Research Bill,” (2006); Yellow rain: Thai bees' Faeces Found. Nature PMID 6709055 (1984); Imamura et al., “Fusarium and Candida Albicans Biofilms on Soft Contact Lenses: Model Development, Influence of Lens Type, and Susceptibility to Lens Care Solutions,” Antimicrob. Agents Chemother. 52(1):171-182 (2008).
Fusarium graminearum commonly infects barley if there is rain late in the season. It is of economic impact to the malting and brewing industries as well as feed barley. Fusarium contamination in barley can result in head blight and in extreme contaminations the barley can appear pink. The genome of this wheat and maize pathogen has been sequenced. Fusarium graminearum can also cause root rot and seedling blight. The total losses in the US of barley and wheat crops between 1991 and 1996 have been estimated at $3 billion.
Rhizoctonia spp. are among the most diverse of plant pathogens, causing root, stem and foliar diseases of many of our most important herbaceous and woody ornamentals. Rhizoctonia spp. usually attack plants at the soil line, causing root loss and constriction of the stem which results in girdling and death of the tops. This pathogen can attack leaves as well and is especially severe when plants are grown close together and kept moist. Entire stock beds or flats can be lost to Rhizoctonia in very short periods of time. The pathogen is soil-borne which means it lives in the soil or potting medium. It causes both pre- and post-emergence damping-off of many ornamental crops such as Vinca, Impatiens, stock, and snapdragon (Chase, A. R., “Rhizoctonia Diseases on Ornamentals,” Western Connection, Turf and Ornamentals (1998)).
Thielaviopsis basicola (Berk. & Br.) Ferraris is a soil inhabitant that attacks more than 100 plant species in 33 families. Members of the Fabaceae, Solanaceae, and Cucurbitaceae families are especially affected by T. basicola (Shew et al., Eds., “Compendium of Tobacco Diseases,”. St. Paul, Minn.: APS Press, pp. 28-29 (1991)). The common name ‘black root rot’ is based on darkly pigmented chlamydospores that form in the root cells of hosts and giving a ‘blackened’ appearance to the root tip (Alexopoulos et al., “Introductory Mycology,” 4th Ed., pp. 869 (1996)). The black root rot fungus is a member of the Hyphomycetes, order Moniliales, family Dematicaceae (Shew et al., Eds., “Compendium of Tobacco Diseases,”. St. Paul, Minn.: APS Press, pp. 28-29 (1991)). General symptoms are root rot and branch dieback. Thielaviopsis basicola can be found in all regions of the world, especially in regions with cool climates. Black root rot can affect a wide range of woody and herbaceous plants including tobacco, holly, begonia, geranium, poinsettia, and pansy (Agrios, G. N., “Plant Pathology,” 4th ed., p. 358 (1997); Alexopoulos et al., “Introductory Mycology,” 4th Ed., pp. 869 (1996); Daughtrey et al., “Compendium of Flowering Potted Plants,” pp. 90 (1995); Lambe et al., “Diseases of Woody Ornamental Plants and Their Control in Nurseries,” pp. 130 (1986); Shew et al., Eds., “Compendium of Tobacco Diseases,” pp. 28-29 (1991)).
Sclerotium rolfsii, an omnivorous, soilborne fungal pathogen, causes disease on a wide range of agricultural and horticultural crops. Although no worldwide compilation of host genera has been published, over 270 host genera have been reported in the United States alone. Susceptible agricultural hosts include sweet potato (Ipomea batatas), pumpkin (Cucurbita pepo L.), corn (Zea mays), wheat (Triticum vulgare) and peanut (Arachis hypogea). Horticultural crops affected by the fungus are included in the genera Narcissus, Iris, Lilium, Zinnia, and Chrysanthemum. See Aycock, R., “Stem Rot and Other Diseases Caused by Sclerotium rolfsii,” N.C. Agr. Expt. St. Tech. Bul., No. 174 (1966); Garren, K. H., “The Stem Rot of Peanuts and its Control,” Virginia Agr. Exp. Sta. Bull. 144 (1959); Paolo, M. A., “A Sclerotium Seed Rot and Seedling Stem Rot of Mango,” Philippine Journal of Science 52:237-261 (1933); Punja, Z. K., “The Biology, Ecology, and Control of Sclerotium rolfsii,” Annual Review of Phytopathology 23:97-127 (1985); Takahashi, T., “A Sclerotium Disease of Larkspur,” Phytopathology 17:239-245 (1927); Townsend et al., “The Development of Sclerotia of Certain Fungi,” Ann. Bot. 21:153-166 (1954); Weber, G. F., “Blight of Carrots Caused by Sclerotium rolfsii, With Geographic Distribution and Host Range of the Fungus,” Phytopathology 21:1129-1140 (1931); Zitter et al., “Compendium of Cucurbit Diseases,” Amer. Phytopath. Soc., St. Paul, Minn. (1966).
Although S. rolfsii is thought to have caused serious crop losses over many centuries, the first unmistakable report of the fungus dates back to 1892 with Peter Henry Rolfs' discovery of the organism in association with tomato blight in Florida. Since Rolfs' report in the late 19th century, the over 2,000 publications on the pathogen support evidence of its worldwide distribution, particularly in tropical and subtropical regions.
The wide host range, prolific growth, and ability to produce sclerotia contribute to the largest economic losses associated with the pathogen. From a global perspective, and local perspective for North Carolina, peanut crops sustain higher losses than any other agricultural crop. In 1959, the United States Department of Agriculture estimated losses from $10 million to $20 million associated with S. rolfsii in the southern peanut-growing region, with yield depletions ranging from 1-60% in fields in the NC coastal plains region.
There exists correlative evidence that certain Trichoderma spp. may be involved in the biological control of several diseases caused by Phytophthora spp., e.g., T viride versus heart rot of pineapple caused by P. parasitica (Papazivas, Ann. Rev. Phytopathol. 23:23-54 (1985)). More compelling correlative evidence is supplied by the well-documented ability of composted hardwood bark (CHB) to provide control of Phytophthora disease of woody plants when incorporated into their rhizosphere (Hoitink et al., Ann. Rev. Phytopathol. 24:93-114 (1986)), including control of crown rot of apple under field conditions (Ellis et al., Plant Dis. 70:24-26 (1986)), and the related documentation that the addition of CHB to a container potting mix resulted in a 100 to 100,000 fold increase in the population levels of T. harzianum in this rooting medium (Nelson et al., Phytopathology 3:1457-1462 (1983)).
Biological control (biocontrol) of plant pathogens is increasingly becoming an essential component in plant disease management. Over-reliance on chemical pesticides, non-sustainable agricultural systems, poor site selection, and resource limitations are examples of agricultural problems faced by growers. Biocontrol offers an alternative to these recurrent/persistent problems in agriculture. Therefore, much emphasis is being placed on the application of such techniques in agriculture.
Many fungi and other microorganisms are known to control various plant pathogens. These biocontrol agents are particularly attractive, because they may be able to protect and colonize plant portions that are particularly inaccessible to conventional agricultural treatments (Harman et al., Seed Sci. and Technol. 11:893-906 (1983)). Trichoderma spp, a filamentous genus of fungi, have been shown to provide varying level of biological control to soil-borne plant pathogens. Five species of Trichoderma are known to be most important for biocontrol. They are T hamatum, T. harzianum, T. konigii, T. polysporum, and T. viride. Desirable and essential traits for biocontrol capability are attributed to specific strains and not the species. For example, strains of T. harzianum have been involved in the treatment of cucumber. While there have been many advances in the use of Trichoderma as a biocontrol agent, it was not until 1992 that this fungus was reported in the treatment of diseases caused by soil-borne Phytophthora spp (Papavizas, Ann. Rev. Phytopathol. 23:23-54 (1985)). Three strains of Gliocladium virens (031, 035, and 041), now known as Trichoderma virens, have been used as biological agents (U.S. Pat. No. 5,165,928 to Smith et al.) to control plant diseases incited by Phytophthora spp, such as root rot, crown, and collar rot (Jeffers et al., Phytopathology 2:533-538 (1982)). However, this invention was limited to the treatment of plant diseases caused by Phytophthora sojae. Additionally, there is the strain GL-21 which is described in U.S. Pat. No. 5,068,105 to Lewis et al. and sold as SoilGard™.
Combinations of different biocontrol agents have been used to control disease. For example, Lewis et al., “A Formulation of Trichoderma and Gliocladium to Reduce Damping-off Caused by Rhizoctonia solani and Saprophytic Growth of the Pathogen in Soiless Mix,” Plant Disease 82:501-06 (1998) uses a formulation of Gliocladium virens TRI-4 and Trichoderma hamatum GL-3, GL-21, or GL-32 for biocontrol. A talc-based formulation known as NUTRI-LIFE TRICHOSHIELD™ has been sold by Nutri-Tech Solutions Pty Ltd. as a plant root growth promoter. This formulation contains a mixture of beneficial fungal species, including Trichoderma harzianum, Trichoderma lignorum, and Gliocladium virens (now Trichoderma virens) together with bio-balancing Bacillus subtilis. Papavizas, et. al., “Effect of Gliocladium and Trichoderma on Damping-off and Blight of Snapbean Caused by Sclerotium rolfsii in the Greenhouse,” Plant Pathology 38: 277-86 (1989) describes the use of 285 wild-type strains and mutants of Gliocladium virens, Trichoderma hamatum, Trichoderma harzianum, and Trichoderma viride against Scelerotium rolfisii in the greenhouse. Ten strains of Gliocladium virens and four strains of Trichoderma harzianum suppressed damping-off of snapbeans by 30-50% and blight by 36-74%. Single strains were as effective as or more effective than mixtures of strains. For instance, the mixture of G1-3 and Th-84 at 3×105 conidia per g soil from each strain was less effective than G1-3 or Th-84 used alone and the triple mixture was least effective. These results suggest to those skilled in the art that Trichoderma harzianum and Gliocladium virens should be used separately to treat plants rather than doing so in combination. In any event, none of the above-described combinations of biocontrol agents involve utilization of a rhizosphere competent Trichoderma harzianum species.
The present invention is directed to overcoming these and other deficiencies in the art.